Effect of cortisol through GR regulated inflammatory cytokine expression and apoptosis of Siberian sturgeon (Acipenser baerii) after LPS treatment

Abstract

AbstractThe various environmental pressures could increase the cortisol levels of sturgeon. However, there is a lack of reports on the mechanism by which elevated cortisol levels affect the immune response of Siberian sturgeon. In this study, a high-level cortisol anti-inflammatory state of Siberian sturgeon after co-treatment with LPS and cortisol was constructed and verified in the primary spleen leukocytes. The glucocorticoid receptor (GR) was cloned and its binding site with Ru486 was analyzed. After adding Ru486 to treat the primary spleen leukocytes, the expression of inflammatory cytokines mediated by GR was explored by qRT-PCR. The structure of spleen after LPS + cortisol injection was observed by histopathology and the changes in apoptotic genes were explored by qRT-PCR. Furthermore, the apoptosis of cells after Ru486 treatment was studied by qRT-PCR, DNA ladder, and Hoechst staining. Firstly, co-treatment with LPS and cortisol led to high levels of serum cortisol and glucose within 24 h after treatment. In this situation, the expression of il-10 and tgf-β1 was upregulated in the LPS + cortisol group, while the expression of il-1β and tnf-α was regulated. However, il-1β, tnf-α, il-10, and tgf-β1 were significantly down-regulated in vitro. Clone analysis of the GR sequence indicated that it is conserved and widely expressed in 18 tissues. Molecular docking simulations suggested that it can bind to cortisol and the GR antagonist Ru486, respectively. Further addition of Ru486 treatment reversed the expression of il-1β, il-10, tgf-β1, and JAK-STAT signaling pathway factors. Histopathological observations showed that LPS + cortisol injection causes splenic cell death. Then, the expression of pro-apoptotic genes p53 and caspase3 increases significantly. After Ru486 treatment in spleen leukocytes, the mRNA expression levels of pro-apoptotic-related genes p53, bax, caspase3, caspase7, and caspase9 were significantly downregulated. GR antagonism could attenuate LPS + cortisol-treated splenic leukocyte DNA fragmentation and nuclear morphological changes. Therefore, cortisol can regulate the production of inflammatory cytokines and promote the process of cell apoptosis through the GR of Siberian sturgeon, thereby inhibiting the inflammatory response.